Nicotine-Induced Effects on Nicotinic Acetylcholine Receptors (nAChRs), Ca2+ and Brain-Derived Neurotrophic Factor (BDNF) in STC-1 Cells

Abstract

In addition to the T2R bitter taste receptors, neuronal nicotinic acetylcholine receptors (nAChRs) have recently been shown to be involved in the bitter taste transduction of nicotine, acetylcholine and ethanol. However, at present it is not clear if nAChRs are expressed in enteroendocrine cells other than beta cells of the pancreas and enterochromaffin cells, and if they play a role in the synthesis and release of neurohumoral peptides. Accordingly, we investigated the expression and functional role of nAChRs in enteroendocrine STC-1 cells. Our studies using RT-PCR, qRT-PCR, immunohistochemical and Western blotting techniques demonstrate that STC-1 cells express several α and β nAChR subunits. Exposing STC-1 cells to nicotine acutely (24h) or chronically (4 days) induced a differential increase in the expression of nAChR subunit mRNA and protein in a dose- and time-dependent fashion. Mecamylamine, a non-selective antagonist of nAChRs, inhibited the nicotine-induced increase in mRNA expression of nAChRs. Exposing STC-1 cells to nicotine increased intracellular Ca2+ in a dose-dependent manner that was inhibited in the presence of mecamylamine or dihydro-β-erythroidine, a α4β2 nAChR antagonist. Brain-derived neurotrophic factor (BDNF) mRNA and protein were detected in STC-1 cells using RT-PCR, specific BDNF antibody, and enzyme-linked immunosorbent assay. Acute nicotine exposure (30 min) decreased the cellular content of BDNF in STC-1 cells. The nicotine-induced decrease in BDNF was inhibited in the presence of mecamylamine. We also detected α3 and β4 mRNA in intestinal mucosal cells and α3 protein expression in intestinal enteroendocrine cells. We conclude that STC-1 cells and intestinal enteroendocrine cells express nAChRs. In STC-1 cells nAChR expression is modulated by exposure to nicotine in a dose- and time-dependent manner. Nicotine interacts with nAChRs and inhibits BDNF expression in STC-1 cells.

Nicotinic acetylcholine receptors (nAChRs) are expressed in Trpm5 positive taste receptor cells (TRCs)

“…Although, the vast majority of the antibodies against nAChR subunits are considered to lack specificity [ 29 , 30 ], in this study nAChR expression in TRCs was confirmed by qRT-PCR, ISH and IHC techniques. Strong supporting evidence for the presence of nAChRs in TRCs is provided by the effect of nAChR agonists and antagonists on the neural and behavioral responses in WT mice, Trpm5 KO mice and rats [ 2 , 3 ] and by our recent studies in STC-1 cells [ 8 ].…”

“…STC-1 cell line is an established cell line of enteroendocrine cells of mouse small intestine. We have recently shown that exposing STC-1 cells to Nic acutely (24 h) or chronically (4 days) induced a differential increase in the expression of nAChR subunit mRNA and protein in a dose- and time-dependent fashion [ 8 ]. Here, we tested, if chronic Nic or ETOH exposure also alters nAChR mRNA expression in CV taste bud cells.…”

“…We have recently [ 8 ] shown that exposing STC-1 cells to 0.25, 0.50, and 1.0 μM Nic acutely (24 h) or chronically (4 days) induced an increase in the mRNA expression of α4, α5, and α6. The changes in α4, α5, and α6 mRNA varied with both Nic concentration and the exposure time.…”

“…The expression of nAChRs has been demonstrated in STC-1 cells [ 8 ] that have been shown to express functional taste receptors involved in the transduction of all five primary taste qualities [ 9 ]. In addition, nAChRs have been shown to be expressed in small intestinal enteroendocrine cells [ 8 ], epithelial cells of intrapulmonary airways that also co-express bitter taste transduction signaling molecules [ 10 ], in bronchial epithelial cells [ 11 ], in colonic mucosal epithelium [ 12 ], in epithelial cells of the small bowel [ 13 ], and human proximal tubular epithelial (HK-2) cells [ 14 ].…”

Nicotinic acetylcholine receptor (CHRN) expression and function in cultured human adult fungiform (HBO) taste cells

“…The changes in FIR were monitored under control conditions and after treating the cells with 0.01 to 1.0 mM nicotine or 10–50 mM ETOH in Ringer’s solution in the absence or presence of 50 μM Mec (Sigma-Aldrich). The FIR values in each cell were normalized to 1 under control conditions [ 7 , 8 , 14 ].…”